The stem cell niche regulates stem cell function through mechanisms not well understood. We have developed an "ex vivo" culture model system, which allows us the unique ability to study stem/progenitor cell behavior response to injury within their native microenvironment. This system was established as a model for the fibrotic lens disease, Posterior Capsule Opacification, which involves collective migration of lens epithelial cells onto the posterior capsule and the emergence of mesenchymal cells. The etiology of this disease is unknown. Recently, a small subpopulation of myo D/G8 antigen positive epiblast-skeletal muscle stem cells has been identified in the lens. In our "ex vivo" model we have identified a these G8 positive (G8pos) epiblast- skeletal muscle stem cells within a niche nestled among the lens epithelial cells of the equatorial zone (EQ), that remain tightly adherent to the lens capsule after our mock cataract surgery. Upon injury in our "ex vivo" model we detect G8 antigen positive epiblast-skeletal muscle stem/progenitor cells moving from their native microenvironment within the EQ zone to the leading edge of the collectively migrating lens epithelial cells. G8pos skeletal muscle stem/progenitor cells may provide the source of 1-smooth muscle actin positive myofibroblasts detected in lens fibrotic diseases. We hypothesize that disruption of the G8pos epiblast- skeletal muscle stem cell niche, such that could occur during lens injury and cataract surgery, causes the aberrant regulation and emergence of mesenchymal cell precursors that 1) drive the collective migration of the lens epithelial cells for wound closure and 2) transdifferentiate into myofibroblasts causing fibrosis. Our "ex vivo" culture model provides the unique opportunity to ask fundamental questions regarding how reciprocal interactions are regulated within the stem cell niche, such as between niche stem/progenitor cells and their surrounding lens cells, and between the cells in the niche and their matrix microenvironment, all in response to wounding. Elucidation of such mechanisms could not only broaden our understanding of the behavior of a subset of stem/progenitor cells in response to injury-induced changes to their niche but also lead to the development of anti-fibrotic therapeutics to preserve vision. The major goals of this proposal are to 1) Examine the hypothesis that G8pos cells found outside the niche following injury to the lens are derived from amplification of the G8pos epiblast-skeletal muscle stem cell population and/or by recruitment of lens epithelial cells to the G8 lineage. 2) Examine the hypothesis that cadherin-mediated cell-cell and integrin-mediated cell- matrix interactions are critical determinants essential to maintain the normal G8 stem cell niche and are altered upon injury for emergence of G8pos mesenchymal precursor cells. 3) Examine the hypothesis that alterations in the G8pos epiblast skeletal muscle stem cell niche causes the aberrant regulation and emergence of mesenchymal precursors that drive collective migration of lens epithelial cells and ultimately transdifferentiate into myofibroblast. PUBLIC HEALTH RELEVANCE: Lens fibrotic disease causes lens opacities leading to the formation of cataracts which block vision. The exact etiology of lens fibrotic disease is unknown and the long term goals of this project are to understand the role of a subpopulation of G8pos skeletal muscle stem cells, which are located in the lens and emerge from their stem cell niche upon injury, in the induction of lens fibrotic disease. Elucidation of such mechanisms could not only broaden our understanding of stem cell behavior in response to injury-induced changes to their niche but also lead to the development of anti-fibrotic therapeutics to preserve vision.